174 KAWASAKI DISEASE SYMPOSIUM ABSTRACTS

95 98 FATAL KAWASAKI DISEASE (KD) DUE TO EARLY FIBROUS OBLITER- A IN LACTOBACILLUS CASEI CELL WALL EXTRACT ATIVE INDUCES CORONARY : AN ANIMAL MODEL FOR KA- Nigel J. Wilson1, Paul Heaton2, Ross Nicholson3, Louise Calder1, Simon Stables4 Paediatric Cardi- WASAKI DISEASEE 1 ology Department, Green Lane Hospital, Auckland, New Zealand , Taranaki Base Hospital, New Trang T. Duong1, Earl D. Silverman2, Martindale V. Bissessar1, Barry L. Myones3, Rae S.M. Yeung2 2 3 Plymouth, New Zealand , Middlemore Hospital, Auckland, New Zealand , Auckland Hospital, Cancer and Blood Program, The Hospital for Sick Children, Toronto, ON, Canada1, Division of 4 Auckland, New Zealand , The Hospital for Sick Children, Toronto, ON, Canada2, Baylor College of Medicine, The purpose of the study is to describe 2 fatal cases of KD due to nonthrombotic obstructive Texas Children’s Medical Center, Houston, TX, USA3 coronary artery disease. Patient 1. 4yr Caucasian male, URTI, low grade , eventually had 4 of Lactobacillus casei cell wall extract (LCWE)-induced coronary arteritis is currently the best animal 5 KD criteria. Given IVIG x 2 about day 15 with no response: persistent and hemolytic model for Kawasaki disease. Kawasaki disease is a multisystem and the leading cause of anaemia 1 month, small joint & 2nd month, abdominal pains 3rd month. Initial acquired disease in children of the developed world. In this study, we show that responses to coronary artery (CA) dilatation progressed with the development of unstable angina and low cardiac LCWE possess all the hallmarks of a superantigen (SAg)-mediated response. LCWE induces massive output 4 months after presentation. Died after anaesthetic induction for cardiac catheterization. Post activation of naive T lymphocytes, as measured by both in vitro proliferation and production of tumor mortem showed thick walled obstructed triple vessel CA disease. Histology showed marked factor-alpha. Interestingly, LCWE-mediated responses require antigen presentation but not fibrocellular intimal proliferation. There was recent myocardial and duodenal . Patient 2. 6 processing by accessory cells. As in the case of other known SAgs, the response to LCWE is MHC month Caucasian male developed fever and 4 criteria for KD but no peripheral oedema or redness. non-classically restricted, and there exists a hierarchy of differing class II MHC in the ability to Given IVIG day 8 and day 20 due to without fever. Echo day 8 and day 50 showed mild present LCWE. The most compelling evidence of superantigenic activity of LCWE is its ability to CA dilatation. He represented day 95 with congestive heart failure, cardiogenic , poor LV induce the characteristic activation and proliferation of T cells expressing specific T cell receptor function and died. Post mortem showed minor CA aneurysmal changes only. There was marked variable beta families 2, 4, 6 and 14, followed by the deletion of these reactive cells. Most luminal obstruction caused by replacement fibrosis of the intimal and media. There was patchy importantly, superantigenic activity of LCWE correlates with its ability to induce coronary vasculitis chronic inflammatory infiltration.In summary, these 2 patients showed atypical KD histology with upon injection into mice. Only LCWE preparations possessing superantigenic properties mediate early intimal fibrosis and luminal obstruction. Conclusions:1. KD may be fatal due to early (within cellular infiltrations in the cardiac leading to coronary arteritis in vivo.Taken together, these 3-4 months from onset) fibrotic obliterative CA disease rather than due to giant CA and findings demonstrate that LCWE contains a novel superantigen, the activity of which may lead to complications.2. New treatment strategies for KD are required for late presenters or non-responders disease pathogenesis. Since LCWE-mediated coronary arteritis in mice is currently the best animal to IVIG when there is evidence of continued disease process. model of Kawasaki disease, understanding of the mechanisms leading from immune activation to coronary artery lesions in this model will be of great benefit in designing new treatment strategies, reducing vascular damage, and, more importantly, the significant morbidity and mortality associated with Kawasaki disease.

96 99 ARTERIAL ANGIOGENESIS IN KAWASAKI DISEASE. A HISTOLOGIC THE ROLE OF T LYMPHOCYTES IN THE PATHOGENESIS OF KA- THREE-DIMENSIONAL RECONSTRUCTIVE STUDY OF A SMALL AR- WASAKI DISEASE IN AN ANIMAL MODEL TERY WITH MANY BRANCHES Wesley C. Chan, Trang D. Duong, Rae S. Yeung Cancer and Blood Research, Hospital for Sick Hirotake Masuda1, Xuefei Hong1, Koichi Kawamura1, Hiroshi Nanjo1, Shiro Naoe2 Second Depart- Children, Toronto, ON, Canada ment of Pathology, Akita University School of Medicine, Akita, Japan1, Department of Pathology, Kawasaki disease (KD) is the most common cause of multisystem vasculitis in children in the Ohashi Hospital, Toho University School of Medicine, Tokyo, Japan2 Anerysm formation with developed world. Although KD is widely believed to be the result of an infectious agent, its etiology inflammatory change is a common pathologic feature of middle sized artery in Kawasaki disease. is still unknown. Past research has reported the possible role for bacterial in the While we have been investigating more than one hundred autopsy cases, we found strange coronary induction of KD. Our laboratory has characterized a novel superantigen found in Lactobacillus casei arterioles of the epicardial space in a case of Kawasaki disease (male, 4 month of age, died in the 38th cell wall extract (LCWE) which is responsible for triggering the immune response leading to KD-like clinical day), which showed many slit-like pits or capillary branchings directly to the capillaries in coronary vasculitis in injected mice. In this study, we investigated the T helper subsets responsible the adventitia. We have never met such lesion in other cases of Kawasaki disease or in other vasculitis for disease induction by studying expression of signature cytokines. LCWE was injected into . To investigate the nature of these lesions, serial three-dimensional histologic reconstruc- susceptible mouse strains and RT-PCR, flow cytometry, and immunohistochemistry were performed tive study of one of the arterioles together with some immunological studies were performed. Within on peripheral lymphoid tissue at set time-points. Expression and production of T helper subset 0.6 mm in length, there are 14 pits and 14 branchings directly to the adventitial capillaries and 2 true associated cytokines were determined. Cardiac tissue was also similarly analyzed to determine the branchings. Luminal surface including pits and branchings is preserved with endothelial layer. There relationship between the immune response in the periphery to that in the heart end organ. Semi- is no inflammatory change. Vascular lesion of Kawasaki disease is characterized by the early vascular quantitation of cytokine mRNA expression demonstrated a Th1 predominated phenotype in the lesion especially on capillaries and later arteritis with formation. However, how small periphery at days 1 to 3 post LCWE injection (IFN␥). This was followed by Th2 cytokine vascular lesions convert to larger arterial lesion especially on aneurysm formation is still mystery. We upregulation after day 3 (IL-4). Flow cytometry confirmed this finding by the detection of intracy- speculate that these lesions are new vascular formation directly from the artery and therefore “arterial toplasmic IFN␥ and IL-4 at similar relative timepoints. In the heart end organ, IFN␥ expression was angiogenesis”. On the point of increase of endothelial surface, angiogenesis and aneurysmal forma- found to be abnormally upregulated 42 days after LCWE injection. Infiltrates identified as T tion are similar, although real features are very different. Vascular lesion of Kawasaki disese should lymphocytes were seen as early as 3 days post LCWE injection and observed in all latter time points be investigated suggesting that there is angiogenesis behind. (up to 6 months). Different T lymphocyte subsets, at different times, are able to mediate the immunologic responses leading to inflammation. Dissecting this immune response to LCWE in mice gives important clues to the etiology of Kawasaki disease in humans.

97 100 PATHOLOGICALLY NORMAL CORONARY ARTERY AFTER KAWASAKI A SWINE MODEL OF PROTEIN MEDIATED VASCULITIS: CLINICAL IM- DISEASE: A CASE REPORT PLICATION FOR KAWASAKI CORONARY ARTERY DISEASE Takashi Fukaya1, Kunizo Baba1, Kei Takahashi2, Shirou Naoe2, Yasuhiko Tomita3, Yasuhiro Ueno4 S Philip1,WCLee1,MHWu3,SKLiu2,HCLue3 Center for Cardiovascular Research, Department Department of , Nishi-Kobe Medical Center, Kobe, Japan1, Department of Pathology, of Comparative Medicine, Pig Research Institute, Miaoli, Taiwan1, Animal Medical Center, New Ohashi Hospital, Toho University School of Medicine, Tokyo, Japan2, Department of Pediatrics, York, USA2, National Taiwan University College of Medicine, Taipei, Taiwan3 Kobe General Hospital, Kobe, Japan3, Department of Legal Medicine, Kobe University School of An attempt was made to produce immune complex vasculitis in piglets, hoping to produce as an Medicine, Kobe, Japan4 experimental Coronary Artery Disease (CAD) animal model by administrating foreign protein, It is well known that the sequelae of the vasculitis are observed pathologically in the coronary mimicking Kawasaki Disease (KD). Systemic type III hypersensitivity reaction by horse serum arteries of the patients with Kawasaki disease. It has been reported that these abnormal findings are injection with clinical implication to CAD in KD was reported only in rabbits. Our study group commonly detected even in patients with clinical diagnosis of normal . We expe- consisted 22 pure bred male piglets of 1.5, 2 and 3 months of age. They were divided into control rienced a case of Kawasaki disease without clinically and pathologically significant coronary artery group (nϭ6) receiving 2 doses of intravenous normal saline, and test group A (nϭ8), receiving 2 lesions. A 1 year 0 month-old male had an episode of Kawasaki disease. The diagnosis was doses at 10 days interval of virus and mycoplasma negative free horse serum (HS), established from the 5 principal clinical findings except . He also had redness of the 2.6gm (5ml) and 5.2gm (10ml) protein/Kg body weight, respectively, test group B (nϭ8) received 3 skin at the site of BCG inoculation. He was treated with intravenous gamma globurin(1g/kg) doses at 5 days interval of HS, 5.2gm protein/Kg body weight. Blood samples were collected for administered two times. Echocardiographic evaluation of the coronary artery was carried out several white blood cells, platelets, vascular endothelial growth factor, liver enzymes, cholesterol and times during the phase of the illness until 2 months after the onset of the disease, and no haptoglobin. Both the test groups developed , echocardiographic coronary artery dilatation and abnormal findings were obtained. His clinical diagnosis was complete recovery from the Kawasaki histopathological changes, whereas none of the control group developed immunopathological re- disease. He died suddenly when he was 2 years 1 month old. He had a febrile episode at night, and sponse. All the animals were sacrificed for the histopathological, and immuno histochemistry study in the next morning, he was found dead. From the macroscopic autopsy findings, the cause of the at different dates after the second HS injection. There were asymmetric inflammatory coronary death was diagnosed as an acute bronchitis. Histologically, the coronary arteries had normal archi- arteritis, such as intimal proliferation, necrosis, vacuolisation, smooth muscle cells proliferation and tecture, and no particular sequelae of arteritis were found. Scar formation was not detected. Mild oedematous changes in arterial walls. The piglets, which received at 5 days interval of 3 doses of HS, intimal thickening was observed in the proximal portion of the bilateral coronary arteries, but it was manifested more skin rashes and histopathological changes in the coronary arteries than those not thought to be pathologically significant. Distal portion of the coronary arteries were normal. received two doses at 10 days interval. Serum sickness is a prototype of immune complex vasculitis, and myocardial ischemia were also excluded. From these findings, we concluded that it is concluded that swine may serve as an experimental model for CAD mimicking KD, for testing there were patients with complete recovery from Kawasaki disease, not only clinically but also the efficacy of pharmacologic therapies. pathologically. This fact may influence the follow up methods of the patients with clinically complete recovery from Kawasaki disease.